Evidence from humans and animals indicate a critical role for the hippocampus in the formation of long- term episodic memories. The hippocampus consists of anatomically segregated subregions with a diversity of neuronal types hypothesized to encode different aspects of an experience. Although the molecular mechanisms of activity-dependent plasticity in hippocampal neurons have been intensely scrutinized, we have limited understanding of the activity patterns shown by select neuronal populations during memory encoding, consolidation and recall. Therefore, there is a strong demand to develop new imaging approaches that can interrogate the activity of hippocampal circuits in the context of the behavior that they are thought to support. Because neuronal spikes are associated with large intracellular calcium conductances, calcium indicators can be used as a proxy for neuronal activation. In the proposed research, we will implement the use of fiber optic fluorescence endomicroscopy (FFE) to chronically monitor the activation of defined hippocampal neurons in mice subjected to hippocampus-dependent tasks. A genetically-encoded calcium indicator will be conditionally expressed in transgenic mice selectively expressing Cre recombinase in CA1 pyramidal cells, or in dentate granule cells and CA3 pyramidal cells. Our initial proof-of-principl experiments in fully awake, unrestricted mice advocate the feasibility of this approach for large-population imaging of single-neuron activity. In our first aim, we will optimize the parameters of FFE required to achieve stable, long-term imaging of the different neuronal populations. In our second aim, we will use this method to define the neuronal ensembles that encode contextual- and trace-fear conditioning, two hippocampus-dependent variants of associative memory hypothesized to engage different circuits. Time-lapse experiments will allow comparisons of the activation patterns observed during training, consolidation and recall. Through genetic or pharmacological interventions against each one of these memory phases, we will link the activity of select neurons to cognition. Thus, the proposed research will advance the field by introducing FFE as a novel in vivo approach to study memory mechanisms, and will use this technique to begin to address previously inaccessible questions on the cellular basis of memory. PUBLIC HEALTH RELEVANCE: Memory impairments are associated with aging and psychiatric diseases. This project uses advanced calcium imaging in behaving mice to reveal the properties of neuronal activation in different types of hippocampal cells during memory formation and retrieval. Consequently, this research may identify specific cellular targets or activity-dependent network patterns that can be exploited to prevent memory impairments.